Angiotensin-converting enzyme inhibitors (ACEIs) are among the most frequently prescribed medications worldwide for the treatment of essential hypertension, left ventricular systolic dysfunction, acute myocardial infarction, and prevention of the progression of diabetic nephropathy. However, the outcome of ACEI treatment varies significantly between individuals and selected populations. Suboptimal response, therapeutic failure, and significant side effects are commonly documented in patients receiving ACEI therapy. Approximately 80% of the ACEIs available for use in the US are synthesized as esterified prodrugs in order to improve otherwise poor oral bioavailability of the active molecule. The activation of ACEI prodrugs primarily occurs in the liver via metabolic de-esterification of th parent drug. The critical activation step is essential in delivering a successful therapeutic outcome since the active metabolites are approximately 10-1000 times more potent relative to their respective parent compounds. Our group and others have demonstrated that carboxylesterase 1 (CES1), the most abundant hydrolase in the liver, is responsible for the activation of ACEI prodrugs in humans. Marked interindividual variability in CES1 expression and activity has been documented, which results in varied therapeutic efficacy and tolerability of many drugs serving as substrates of CES1. Genetic variation of CES1 is considered to be a major factor contributing to variability in CES1 function. We are among the first demonstrating that some CES1 single nucleotide polymorphisms (SNPs), such as the G143E, are loss-of- function variants, which can significantly affect both pharmacokinetics (PK) and pharmacodynamics (PD) of medications metabolized by CES1. In this proposal, we will test the hypothesis that CES1 genetic variation contributes to interindividual variability of CES1 function, and is one of the determinants influencing the activation and subsequent pharmacological activity of ACEI prodrugs. Specific Aim 1 is to conduct a multiple- dose healthy volunteer study to evaluate the impact of CES1 genetic variation on the activation, PK, and PD of enalapril, a model ACEI prodrug activated by CES1. Specific Aim 2 is to identify and characterize functional coding and regulatory CES1 variants that affect the activity and expression of CES1 using several complementary in vitro approaches. The achievement of these stated aims will represent a major step towards the establishment of an evidence base from which a more individualized use of ACEI prodrugs can emerge. This project also has significant potential for the improvement of therapeutic efficacy and safety of many other medications metabolized (activated or deactivated) by CES1.